| Co-Investigator(Kenkyū-buntansha) |
ENDO-ICHIKAWA Yoko Kansai Medical University, Public Health, Assistant Professor, 医学部, 講師 (50193438)
KOHNO Hirao Kansai Medical University, Hygiene, Assistant Professor, 医学部, 講師 (30148522)
TAKETANI Shigeru Kansai Medical University, Hygiene, Associate Professor, 医学部, 助教授 (20121949)
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| Research Abstract |
We examined formation of DNA adducts or induction of DNA-damage responsive gene by exposure of yeast expressing mammalian cytochrome P-450 isozymes to chemical carcinogens. The achievement of these systems will facilitate to simply evaluate the chemical carcinogens. When yeast exhibiting rat and human cytochrome P-450 1A1,1A4 and 2B5 was exposed to carcinogens such as alfatoxin B1,2-aminofluorene and 2-acethylaminofluorene (AAF), DNA-adducts in yeast chromosomes were detected, but these structures were different among chemicals. RNR3, a small subunit of ribonucleotide reductase, is one of potent DNA-damage responsive genes. RNR3mRNA in yeast was markely induced by the treatment with DNA damage chemicals, including 4-nitroquinoline-1-oxide, cysplatin and bleomycin. Then the 5'-promoter region of RNR3 was connected to the structural gene of beta-galactosidase, and the reporter assay was made. Exposure of yeast to aflatoxin and AAF resulted in the induction of reporter activity, and these inductions depend on the expression of mammalian P-450. Therefore, construction of yeast expressing various P-450s will be useful to detect carcinogenic potency.
To clarify cis-acting regulation of RNR3 gene by DNA damage, deletion analysis of the promoter region was performed. Then we found that at least three potential cis-elements are neccessary for the inducible expression of RNR3 in response to DNA damage. Finally we examined induction of mammalian DNA-damage responsive genes in cultured cells, and showed that several genes such as gadd152 and gadd74 were induced by exposure to carcinogens. However, the mechanisms involved in the inductions are complicate or we cannot figure out the regulations of mammalian gene expression in response to DNA damage.
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